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Ammonium sulphate precipitation of recombinant adenovirus from culture medium: an easy method to increase the total virus yield

Gene Therapy volume 7, pages 1570–1574 (2000)Cite this article

Abstract

In the majority of the methods for purifying and concentrating recombinant adenoviruses (rAds) the virus that is associated with the helper cells is harvested, while the virus that is present in the cell-culture medium is discarded. During routine propagation of adenovirus type-5 vectors at optimised conditions we noted that, on average, 47% of the total amount of virus is present in the culture medium. To recover and concentrate these rAds from the medium, we devised a method, which is based on ammonium sulphate ((NH4)2SO4) precipitation. At 40% (NH4)2SO4 saturation, 95 ± 6% of the available virus precipitates from the medium, while the majority of the protein (85%) remains in solution. In contrast to adenovirus precipitation with polyethylene glycol, the (NH4)2SO4precipitation technique allows collection of precipitated rAds by filtration. We demonstrate here that (NH4)2SO4 precipitation of rAds from cell-culture medium is a simple and fast technique that can be used in combination with standard virus isolation methods to increase the yields of rAds.

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Acknowledgements

We would like to thank Anneke Goedheer (IntroGene, Leiden) for the performance of the HPLC analysis and Steve Cramer and Diana van den Wollenberg for their excellent technical assistance. This work was financially supported by grants from the Dutch Organisation for Scientific Research (NWO), project No. 901–01–096.

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  1. Department of Molecular Cell Biology, Leiden University Medical Centre (LUMC), Wassenaarseweg 72, Leiden, 2333 AL, The Netherlands

    F H E Schagen, H J Rademaker, M J W E Rabelink, H van Ormondt, F J Fallaux, A J van der Eb & R C Hoeben

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  1. F H E Schagen
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Schagen, F., Rademaker, H., Rabelink, M. et al. Ammonium sulphate precipitation of recombinant adenovirus from culture medium: an easy method to increase the total virus yield. Gene Ther 7, 1570–1574 (2000). https://doi.org/10.1038/sj.gt.3301285

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